Pathogenic mechanisms underlying late phase reactions (LPR) in IgE- mediated responses in skin and respiratory tract are not well- defined with roles postulated for activated granulocytes and T lymphocytes. Using skin chamber approach overlying developing LPR to pollen Ag challenge, we have made frequent sequential assessments, finding characteristic sequential patterns of mast cell, (MC) and likely basophil, mediator release, accumulation of activated neutrophils and eosinophils, increased expression of adhesion molecules on granulocytes and vascular endothelium, followed by entry of CD4+ lymphocytes. However, it is still unclear what initiates these vascular and granulocyte responses and their relationship to the pathogenesis of LPR. We will extend these studies in a group of inter-related projects to learn more about mechanisms underlying these inflammatory events. Using a combination of skin chamber and skin biopsy approaches, we will: 1) compare patterns of PMN and eosinophil activation, vascular adhesion molecule expression, mediator release and the presence of cytokine mRNA and protein in different subjects with and without LPR developing after similar immediate responses (looking for a host factor in LPR); 2) determine why the same subject manifest strong LPR to pollen antigens why codeine induces strong immediate MC activation and whealing but nosubsequent LPR in the sites; 3) assess adherence of LPR site granulocytes to matrix proteins; since such adherence is likely important in leukocyte passage from the blood into and through the dermis 4) determine the possible role of T lymphocytes in LPR, - whether there is preferential reactivity of lymphocytes from LPR sites to the challenge Ag, by Vbeta gene usage and limiting dilution approaches; also whether this associated with certain cytokine profiles; 5) study mechanisms underlying corticosteroid inhibition of LPR. These studies will utilize immunoassays, immunohistochemistry, electron microscopy, cell culture and molecular biology approaches. The overall hypothesis being tested is that events following initial MC activation, including the continued presence of antigen in the site, lead to release of mediators and cytokines that upregulate accumulation and activation of granulocytes in the site. These events may lead to subsequent accumulation of T cells, which perpetuate the reaction through cytokine release. Corticosteroids may inhibit LPR by suppressing cytokine formation and granulocyte adhesion/activation. These findings will be considerable clinical important because the inflammatory response in LPR is thought to play a major role in chronic allergic diseases.